Final Report Summary - MINICELL (Building minimal cells to understand active cell shape control)
Stand alone description of the project and its outcomes
The aim of this ERC project was to understand the physical mechanisms that enable animal cells to control their shape. Cells combine a high resilience against mechanical stress with the ability to change shape during processes such as cell division and migration. This paradoxical combination of strength and deformability depends on the cell cortex, a thin meshwork of actin filaments right underneath the cell membrane. This cortex provides strength and rigidity because it is crosslinked by specialized linker proteins and it also drives cell shape changes by means of molecular motors that actively generate contractile forces. It is important to understand how cells regulate the mechanical behavior of this so-called actin cortex because of its central role in many physiological functions (such as embryogenesis and tissue repair) and in diseases (such as cancer). However, it is challenging to understand the molecular basis of cell shape regulation due to the enormous molecular complexity of cells. Therefore, the approach of this ERC project was to construct simplified, synthetic cells by combining model biomembranes with a model actin cortex built from purified cellular constituents. This bottom-up approach is ideally suited to understand how cell-scale behavior emerges from molecular interactions and to provide input for quantitative theoretical models of cell mechanics. The main outcomes of the project were the following:
1) Regulation of cortex-membrane adhesion: We used in vitro reconstitution to define the molecular mechanisms by which septins mediate anchoring of the actin cortex to the cell membrane. Septins play an essential role in cell shape control in all animals, but the underlying mechanisms remain poorly understood. We established a toolset of imaging modalities and quantitative surface analytical tools to study the interactions of septins with lipid membranes and actin filaments in synthetic cell model systems. We discovered that septins strongly interact with lipid membranes through electrostatic interactions with anionic lipids and assemble into dense filamentous scaffolds that provide membrane rigidity. We furthermore discovered that septins can anchor actin-myosin networks to membranes and regulate myosin-driven activity.
2) Cell shape control by the cortex: We used rheological measurements and computational modelling to study how the actin cortex is able to combine mechanical strength with deformability. We discovered that an important factor is the fact that actin filaments are transiently connected by linker proteins. These cause elastic rigidity on short-timescales, whilst allowing for viscoelastic flows on long timescales. We showed that transient crosslinks make actin networks vulnerable to mechanical stress because crosslink unbinding under force leads to spontaneous network fracture. However, special crosslinks that exhibit catch bond behavior can prevent network fracture while ensuring high network deformability.
3) Cell shape polarization by cortical contractility: We combined time-lapse imaging with computer simulations to study how molecular interactions between actin filaments and myosin-2 motors translate into active cortex contraction. We discovered that myosin motors mediate polarity sorting of actin filaments, which eventually causes network contraction. We developed a unique experimental setup that combines a confocal scanning unit with various laser-based micromanipulation tools, which allows us to study the mechanics and contractility of actin cortical networks inside cell-sized vesicles.
The aim of this ERC project was to understand the physical mechanisms that enable animal cells to control their shape. Cells combine a high resilience against mechanical stress with the ability to change shape during processes such as cell division and migration. This paradoxical combination of strength and deformability depends on the cell cortex, a thin meshwork of actin filaments right underneath the cell membrane. This cortex provides strength and rigidity because it is crosslinked by specialized linker proteins and it also drives cell shape changes by means of molecular motors that actively generate contractile forces. It is important to understand how cells regulate the mechanical behavior of this so-called actin cortex because of its central role in many physiological functions (such as embryogenesis and tissue repair) and in diseases (such as cancer). However, it is challenging to understand the molecular basis of cell shape regulation due to the enormous molecular complexity of cells. Therefore, the approach of this ERC project was to construct simplified, synthetic cells by combining model biomembranes with a model actin cortex built from purified cellular constituents. This bottom-up approach is ideally suited to understand how cell-scale behavior emerges from molecular interactions and to provide input for quantitative theoretical models of cell mechanics. The main outcomes of the project were the following:
1) Regulation of cortex-membrane adhesion: We used in vitro reconstitution to define the molecular mechanisms by which septins mediate anchoring of the actin cortex to the cell membrane. Septins play an essential role in cell shape control in all animals, but the underlying mechanisms remain poorly understood. We established a toolset of imaging modalities and quantitative surface analytical tools to study the interactions of septins with lipid membranes and actin filaments in synthetic cell model systems. We discovered that septins strongly interact with lipid membranes through electrostatic interactions with anionic lipids and assemble into dense filamentous scaffolds that provide membrane rigidity. We furthermore discovered that septins can anchor actin-myosin networks to membranes and regulate myosin-driven activity.
2) Cell shape control by the cortex: We used rheological measurements and computational modelling to study how the actin cortex is able to combine mechanical strength with deformability. We discovered that an important factor is the fact that actin filaments are transiently connected by linker proteins. These cause elastic rigidity on short-timescales, whilst allowing for viscoelastic flows on long timescales. We showed that transient crosslinks make actin networks vulnerable to mechanical stress because crosslink unbinding under force leads to spontaneous network fracture. However, special crosslinks that exhibit catch bond behavior can prevent network fracture while ensuring high network deformability.
3) Cell shape polarization by cortical contractility: We combined time-lapse imaging with computer simulations to study how molecular interactions between actin filaments and myosin-2 motors translate into active cortex contraction. We discovered that myosin motors mediate polarity sorting of actin filaments, which eventually causes network contraction. We developed a unique experimental setup that combines a confocal scanning unit with various laser-based micromanipulation tools, which allows us to study the mechanics and contractility of actin cortical networks inside cell-sized vesicles.